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Preliminary Study of in vivo electroporation of VEGF-C Gene Transfer for Lymphedema Model

体内电穿孔VEGF-C基因转染淋巴水肿模型的初步研究

基本信息

批准号：
15591365
负责人：
AMANO SADAO
金额：
$ 2.24万
依托单位：
NIHON UNIVERSITY
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2003
资助国家：
日本
起止时间：
2003 至 2004
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-15591365/
关键词：
VEGF-C Lymphedema Lymphangiogenesis Gene transfer post operative state of breast cancer in vivo electroporation

项目摘要

We have examined the relationship between the vascular endothelial growth factors (VEGE-family) and the character of cancer (Mori, Saitou, Kuboi, et al.). On the other hand, the patients who received axillary dissection and postoperative irradiation for the breast cancer produce the lymphedema of the suffered arm that are difficult to cure (Kitajima et al.). We conducted the experimental study of the gene transfer of VEGF-C (lymphatic duct growth factor) to determine the effect and safety of this therapy for lymphatic edema animal model.Method : The pCAGGS-VEGF-C expression plasmid was used to assess the efficiency of gene transfer into muscle by electroporation in vivo. This vector was prepared by inserting a VEGF-C cDNA into the pCAGGS expression vector containing the CAG promoter. In vivo electroporation method was performed using the gene transfer equipment CUY21EDIT. The surgical lymphedema by the method of Kawashima was created in the leg of one side of the rat. The rat were injected with 50 μg each of closed circular plasmid DNA (pCAGGS-VEGF-C, control pCAGGS). Circumference measurement was performed to calculated percent difference and circumference reduction rate. The tibialis anterior muscles samples for sacrificed rat were fixed 10% formalin in PBS for 24hours. Serial cut sections were sliced for hematoxylin-eosin (H-E) and immunohistochemical staining to evaluate lymphatic vessel findings (vessel density and vessel area rate) histologically. Results : The percent difference and the circumference reduction rate respectively showed between VEGF group and control. However, a significant difference was not found between the treatment group and control group. Lymphatic vessels density and vessel area rate were also increased in treatment group however ; there was no significant difference between the two groups. Conclusion : In vivo electroporation gene transfer of VEGF-C cDNA was demonstrated likelihood of effective to the lymphedema.

我们研究了血管内皮生长因子（VEGE家族）与癌症特征之间的关系（Mori、Saitou、Kuboi等人）。另一方面，接受腋窝淋巴结清扫和乳腺癌术后放射治疗的患者会产生难以治愈的患臂水肿（Kitajima等人）。为探讨VEGF-C（lymphatic duct growth factor，淋巴管生长因子）基因转染治疗淋巴水肿动物模型的有效性和安全性，本研究采用电穿孔法将pCAGGS-VEGF-C表达质粒导入肌肉，观察其对淋巴水肿动物模型的治疗效果。通过将VEGF-C cDNA插入到含有CAG启动子的pCAGGS表达载体中来制备该载体。使用基因转移设备CUY 21 EDIT进行体内电穿孔方法。通过Kawashima的方法在大鼠一侧的腿中产生手术性水肿。注射闭环质粒DNA（pCAGGS-VEGF-C，对照pCAGGS）各50 μg。进行周长测量以计算百分比差异和周长减少率。将处死大鼠的胫骨前肌样本用PBS中的10%福尔马林固定24小时。连续切片进行苏木精-伊红（H-E）和免疫组织化学染色，以评价组织学上的淋巴管发现（血管密度和血管面积率）。结果：VEGF组与对照组比较，差异有显著性意义（P <0.05），VEGF组与对照组比较，差异有显著性意义（P <0.05）.但治疗组与对照组之间无显著性差异。治疗组淋巴管密度和淋巴管面积率也有所增加，但两组间无显著性差异。结论：电穿孔法将VEGF-CcDNA基因导入小鼠体内，可有效治疗小鼠水肿。